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Biomedical subjects

D H Boldt

Publications and source records attributed to D H Boldt.

68 records · Page 4Linked to original sources
PubMed1980

Fractionation of human lymphocytes with plant lectins. III. Identification of cells regulating the in vitro response to L-phytohaemagglutinin.

Peripheral blood lymphocytes (PBL) were separated into two subclasses by differential adherence to wheat germ agglutinin (WGA). WGA-adherent PBL differed structurally (different WGA-binding properties) and functionally from WGA-non-adherent cells. As judged by [3H]-thymidine incorporation, WGA-adherent PBL responded less well to L-phytohaemagglutinin (L-PHA) than non-adherent cells. This difference was not due to different L-PHA dose requirements or different response kinetics. WGA-adherent and non-adherent PBL bound identical amounts of 125I-L-PHA and contained comparable percentages of T cells, B cells, and monocytes. Addition of mitomycin-C-pre-treated WGA-adherent cells to non-adherent cells caused suppression of the L-PHA response. Maximal suppression occurred at a ratio of 1 adherent:2 non-adherent cells and on days 5-7 of culture. Under these conditions the adherent cells themselves did not proliferate indicating that active proliferation was not required for inhibition. Suppression was selective for L-PHA as it did not occur in cultures stimulated with concanavalin A, pokeweed mitogen, Lens culinaris lectin, or in the mixed leucocyte reaction. Cell fractionation techniques indicated that plastic adherent cells (presumably monocytes) in the WGA-adherent subclass were critical for mediation of suppression; These data provide evidence for a specific human suppressor cell of the in vitro response to L-PHA.

Cell Adhesion↗
PubMed1978

Different effects of concanavalin A and E-phytohemagglutinin upon lymphocyte glycosyltransferase activities.

We have studied glycosyltransferase activities in human lymphocytes stimulated with the plant mitogens E-phytohemagglutinin or concanavalin A (Con A) and have compared the results with activities found in resting lymphocytes. Compared to resting lymphocytes, Con A-stimulated lymphocytes possess an enhanced capacity to transfer the sugars, sialic acid, galactose, and N-acetyl-glucosamine from their respective nucleotide donors to both endogenous cellular acceptors and added exogenous glycoprotein acceptors. The enhanced glycosyltransferase activity induced by Con A is not inhibited by puromycin despite effective inhibition of de novo protein synthesis, indicating that synthesis of new glycosyltransferase enzymes is not necessary for the observed increases in glycosylation activity. By contrast to the findings in Con A-stimulated cells, the corresponding glycosyltransferase activities of E-phytohemagglutinin-stimulated lymphocytes do not differ from those of unstimulated lymphocytes. These data indicate that individual plant lectins have different effects on the biosynthesis of complex saccharides by cultured human lymphocytes.

Cell Membrane↗
PubMed1977

Leukemic reticuloendotheliosis: "hairy cell leukemia," functional and structural features of the abnormal cell in a patient with profound leukocytosis.

The development of profound leukocytosis in a patient with leukemic reticuloendotheliosis (LRE) enabled us to obtain purified LRE cells for the investigation of their structural and functional characteristics. The LRE cells of our patient bore surface immunoglobulin and had complement receptors but did not bear Fc receptors and did not form rosettes with sheep erythrocytes. By electron microscopy, the cells were observed to contain typical ribosome lamella structures and to phagocytize both 0.81 micron latex particles and complement-coated zymosan particles. They were adherent to both glass and nylon wool fibers. The mitogenic response to erythroagglutinating phytohemagglutinin was normal to magnitude but delayed chronologically. The binding of 125I-labeled plant lectins was used to characterize the surface topography of LRE cells. Results of these studies indicated that the LRE cell surface differed significantly from the surface of normal T and B lymphocytes and chronic lymphatic leukemia cells. The LRE cells were capable of both stimulating and responding in a one-way mixed lymphocyte culture. However, the LRE cells were not active as effector cells of either cell-mediated lympholysis, a T cell function, or antibody-dependent cellular cytotoxicity, a null cell function. In contrast, they were effector cells of lectin-induced cellular cytotoxicity showing that they did possess the capacity to function as cytotoxic effector cells. These data indicated that the LRE cells in our patient had surface and functional characteristics of both lymphocytes and monocytes.

Binding Sites↗
PubMed1977

Excretion of DNA by purified human lymphocyte subpopulations.

A large proportion of DNA synthesized in vitro by human lymphocytes stimulated with plant mitogens or specific antigens is selectively excreted from the cells. To determine if DNA excretion differs among various types of lymphocytes, we examined purified human lymphocyte subpopulations for DNA synthesis and excretion in response to stimulation by L-PHA. The relative proportion of newly synthesized DNA that is excreted by unseparated mononuclear cells, macrophage-depleted cells, T, and B lymphocytes is identical despite great differences in the magnitude of their responses. Low levels of both DNA synthesis and excretion by macrophage-depleted cells and B cells can be increased by reconstitution with macrophages and T cells, respectively. These data indicate that DNA exretion is a general property of lymphocytes stimulated to undergo DNA synthesis by plant mitogens.

Antigens↗
PubMed1976

Rosette formation between human lymphocytes and sheep erythrocytes. Inhibition of rosette formation by specific glycopeptides.

Rosette formation with unsensitized sheep erythrocytes is a characteristic of human thymus dependent lymphocytes. Release of glycopeptides from the sheep erythrocyte by trypsin reduces rosette formation. These tryptic glycopeptides inhibit rosette formation by untrypsinized sheep erythrocytes; this suggests that rosetting is mediated by erythrocyte surface glycopeptides. To investigate the molecular nature of this interaction, we examined the abilities of various model compounds to act as haptenic inhibitors of rosette formation. Inhibition is given by glycopeptides bearing oligosaccharide units rich in sialic acid, galactose, N-acetylglucosamine, and mannose linked to asparagine residues through glycosylamine bonds. Among compounds tested, fetuin glycopeptide is most effective, but human transferrin glycopeptide and human erythrocyte glycopeptide I also inhibit rosette formation. Other compounds including human erythrocyte glycopeptide II, human IgG glycopeptide, lacto-N-neotetraose, 3'- and 6'-sialyllactose show no significant inhibition. Neither sialic acid, galactose, manose, nor N-acetyl-glucosamine alone inhibits rosette formation. Stepwise degradation of fetuin glycopeptide established the galactose residues as important determinants of inhibitory activity. Fetuin glycopeptide blocks rosette formation when added to a suspension of human lymphocytes and sheep erythrocytes or when preincubated with human lymphocytes, but not when preincubated with sheep erythrocytes. Studies of the binding of [3H] fetuin glycopeptide to normal lymphocytes demonstrate 7.5 x 10(6) saturable binding sites per cell. No saturable binding of this compound to sheep erythrocyte membranes is observed. Compared to normals, lymphocytes from patients with chronic lymphatic leukemia demonstrate decreased fetuin glycopeptide binding with a mean of 0.9 x 10(6) sites per cell. This decreased binding correlates with the impaired ability of these cells to form rosettes. The data suggest that fetuin glycopeptide inhibits rosette formation by binding to the thymus-dependent cell where competition occurs with sheep erythrocytes for specific lymphocyte surface receptors.

Animals↗
PubMed1975

Interaction of plant lectins with purified human lymphocyte populations: binding characteristics and kinetics of proliferation.

With six purified plant lectins as probes of cell surface structure, human T, B, and Null lymphocytes have been examined for cell surface differences. The results of binding experiments with radioiodinated lectins indicate that these three lymphocyte populations, although functionally different, bind equivalent amounts of E- and L-PHA, lentil PHA, Con A, RCA-I, and WGA. Since these lectins are capable of binding to at least four different cell surface oligosaccharides, these data indicate that human T, B, and Null lymphocytes do not differ with respect to surface content of these carbohydrate structures. In addition, all six lectins were investigated for their ability to stimulate DNA synthesis in these human lymphocyte populations. Four of these lectins (E- and L-PHA, lentil PHA, and Con A) were mitogenic for human lymphocytes, whereas RCA-I and WGA failed to stimulate significant DNA synthesis in any lymphocyte population. When incubated with the mitogenic lectins, both B and Null cells displayed a delayed peak of DNA synthesis in comparison to T cells. Thus, despite virtually identical binding characteristics, the lectin-induced mitogenic response exhibited by B and Null cells was markedly different kinetically from that of T cells.

Adsorption↗
PubMedDate not supplied

Expression of different NF-kappaB pathway genes in dendritic cells (DCs) or macrophages assessed by gene expression profiling.

NF-kappaB/Rel transcription factors have been implicated in the differentiation of monocytes to either dendritic cells (DCs) or macrophages, as well as in the maturation of DCs from antigen-processing to antigen-presenting cells. Recent studies of the expression pattern of Rel proteins and their inhibitors (IkappaBs) suggest that their regulation during this differentiation process is transcriptional. To investigate differential gene expression between macrophages and DCs, we used commercially available gene microarrays (GEArray KIT), which included four of the NF-kappaB/Rel family genes (p50/p105, p52/p100, RelB, and c-rel) and 32 additional genes either in the NF-kappaB signal transduction pathway or under transcriptional control of NF-kappaB/Rel factors. To generate macrophages and DCs, human adherent peripheral blood monocytes were cultured with M-CSF or GM-CSF + IL-4 respectively for up to 8 days. DCs (and in some experiments, macrophages) were treated with lipopolysaccharide (LPS) for the last 48 h of culture to induce maturation. Cells were harvested after 7 days, cDNA was prepared and radiolabeled with alpha-(32)P-dCTP, then hybridized to gene arrays containing specific gene probes. beta-actin and GAPDH or PUC18 oligonucleotides served as positive or negative controls, respectively. The expression of all four NF-kappaB/Rel family genes examined was significantly upregulated in maturing DCs compared to macrophages. The strongest difference was observed for c-rel. RT-PCR determinations of c-rel, RelB, and p105 mRNAs confirmed these observations. Among the 32 NF-kappaB/Rel pathway genes, 14 were upregulated in mature DCs compared to macrophages. These genes were IkappaBalpha, IKK-beta, NIK, ICAM-1, P-selectin, E-selectin, TNF-alpha, TNFR2, TNFAIP3, IL-1alpha, IL-1R1, IL-1R2, IRAK, and TANK. By contrast, only mcp-1 (monocyte chemotactic protein 1) was upregulated in macrophages compared to DCs. NF-kappaB pathway genes upregulated in DCs compared to macrophages were constitutively expressed in monocytes then selectively downregulated during macrophage but not DC differentiation. LPS did not induce expression of most of these genes in macrophages but LPS did induce upregulation of IL-8 in mature macrophages. We conclude that NF-kappaB/Rel family genes, especially c-rel, are selectively expressed during differentiation of monocytes towards DCs. Moreover, this differential expression is associated both with activation of different NF-kappaB signal transduction pathways in DCs and macrophages and with expression of a unique subset of genes in DCs that are transcriptionally targeted by NF-kappaB/Rel factors. The results illustrate the ability of the NF-kappaB pathway to respond to differentiation stimuli by activating in a cell-specific manner unique signalling pathways and subsets of NF-kappaB target genes.

Cell Differentiation↗
PubMedDate not supplied

A phase I clinical and pharmacological profile of dacarbazine with autologous bone marrow transplantation in patients with solid tumors.

Dacarbazine (DTIC) is a chemotherapy drug which has antitumor activity at standard doses, exhibits a steep dose-response effect in vitro, and is associated with relatively few non-hematologic toxicities. These characteristics suggest a potential role for this drug in bone marrow transplant preparative regimens. To pursue this hypothesis, 16 patients with refractory solid tumors were enrolled in a phase I study of single agent DTIC to determine the dose of DTIC requiring bone marrow reinfusion and to define the dose-limiting toxicity and maximum tolerated dose when given with autologous bone marrow rescue. Pharmacokinetics were evaluated at the 4394 mg/m2 dose level. The marrow requiring dose was 2000 mg/m2 when given as a single intravenous (i.v.) infusion. The extramyeloid dose-limiting toxicity of DTIC was hypotension, with the maximum tolerated dose of DTIC being 3380 mg/m2 when given with bone marrow transplantation (BMT). Other toxicities were transient and tolerable. At 4394 mg/m2 of DTIC, plasma concentrations declined biexponentially with a terminal half-life of 3 hours. The mean clearance was 10.6 L/hr/m2 with a volume of distribution at steady state of 37.5 L/m2 and a mean maximum plasma concentration of 150 mcg/ml. One patient with melanoma developed a partial response of short duration after receiving 2600 mg/m2 of DTIC. Dacarbazine can be significantly dose escalated with an acceptable toxicity profile, when given with BMT. Future trials should focus on the addition of this drug to current BMT preparative regimens used for the treatment of patients with lymphoma.

Adult↗
PubMedDate not supplied

Cellular immunology in the baboon (Papio cynocephalus): examination of structural and functional characteristics of adult and fetal lymphoid cells.

We studied structural and functional characteristics of lymphocytes from adult and fetal baboons (Papio cynocephalus). Flow cytometry with monoclonal antibodies to human lymphocyte antigens and plant lectins was used to define expression of surface antigens on lymphocytes from adult and 140 day fetal baboons (term = 180 days). Major T cell antigenic determinants on adult and fetal baboon lymphocytes were the Tp50, Tp32-45, and p45 glycoproteins detected by monoclonal reagents T11, OKT8, and OKT10 respectively. Baboon T lymphocytes did not react with the OKT3/anti-Leu4 or OKT4/anti-Leu3a reagents which detect, respectively, Tp19-29 and Tp55, major surface glycoproteins on human T lymphocytes. OKT6, which identifies the human TL antigen equivalent on thymocytes, did not react with baboon thymocytes. These data demonstrate major evolutionary divergence between human and baboon T lymphocytes. By contrast, baboon lymphocytes resembled human peripheral lymphocytes in reactivities with several non-T cell reagents. Lectin binding studies revealed substantially fewer peanut agglutinin-and wheat germ agglutinin-binding cells in suspensions of baboon fetal splenocytes and adult peripheral lymphocytes compared with fetal thymocytes. Therefore, maturation of baboon T lymphocytes is associated with loss of surface carbohydrate structures that bind these lectins. Adult and fetal baboon lymphocytes resembled human and murine lymphocytes in their capabilities to respond to mitogens and to produce interleukin-2. As in other species, adult, but not fetal baboon lymphocytes, mediated NK activity against a variety of nucleated target cells. Despite divergence in lymphocyte antigen expression, baboon lymphocyte functional development closely parallels that seen in humans.

Animals↗